Abstract Objectives: Alcohol is a known breast cancer risk factor and it is also associated with increased mammographic breast density, a well-established, strong breast cancer risk factor. Despite some evidence on the associations of alcohol use with tissue composition on mammograms, no studies have examined these associations using direct measurement of tissue composition in normal breast tissue of cancer-free women. We investigated the associations of recent and cumulative average alcohol use with percentage of epithelium, stroma, fibroglandular (epithelium+stroma), and fat tissue in benign breast biopsy samples. Methods: This study included 859 cancer-free women with biopsy-confirmed benign breast disease (BBD) within the Nurses’ Health Study and Nurses’ Health Study II cohorts. Percentage of each tissue type was measured on whole section images with a deep-learning technique. All tissue measures were log-transformed. The data on alcohol use was obtained from semi-quantitative food frequency questionnaires. Information on other covariates was collected on initial questionnaire and updated biennially thereafter. We examined associations for recent (at the time of biopsy) and cumulative average alcohol use (from all questionnaire preceding the biopsy), both of which were modeled as a continuous (g/day) and categorical (using various approaches: 0, 0-<5, and ≥5 g/day; non-drinkers, <11 g/day [<1 drink/day], 11-<22g/day [1-<2 drinks/day], and ≥ 22 g/day [≥2 dinks/day]; and quartiles based on the distribution in the study sample). Generalized linear regression was used to examine the associations of alcohol with % of tissue types, while adjusting for known breast cancer risk factors and BBD subtype. Results: In this study of 859 cancer-free women, 260 (30.3%) had non-proliferative disease, 485 (56.5%) had proliferative disease without atypia, and 114 (13.3%) had atypical hyperplasia, consistent with previously reported distributions of these BBD subtypes. In our study, 30.3% of women consumed ≥5 g/day of alcohol at the time of biopsy and 18.1% consumed ≥11 g (≥1 drinks) of alcohol per day. The average % of epithelium, stroma, and fat in our study was 9.0% (range 0.7-52.2%), 72.4% (range 23.6-99.0%), and 18.6% (range 0-71.3%), respectively. Both recent and cumulative alcohol use were inversely associated with % of stroma and fibroglandular tissue; these associations were most pronounced for recent use of ≥22 g (≥2 drinks) per day (stroma: β=-0.08, 95% Confidence Interval [CI] -0.13;-0.03; fibroglandular: β=-0.09, 95% CI -0.13;-0.04) and cumulative average use of ≥22 g (≥2 drinks) per day (stroma β=-0.08, 95% CI -0.13;-0.02; fibroglandular: β=-0.09, 95% CI -0.14;-0.04). Recent and cumulative average use of ≥22 g/day were also positively associated with % of fat (β=0.30, 95% CI 0.03; 0.57 for recent and β=0.32, 95% CI 0.04; 0.61 for cumulative average). In stratified analysis by menopausal status, recent and cumulative average alcohol use were inversely associated with % of stroma and fibroglandular tissue and positively associated with % of fat in postmenopausal women. The strongest associations were observed for recent alcohol use of ≥22 g/day (stroma: β=-0.12, 95% CI -0.21;-0.04; fat: β=0.67, 95% CI 0.23;1.12; fibroglandular: β=-0.15, 95% CI -0.23;-0.07) and cumulative average use of ≥22 g/day (stroma β=-0.14, 95% CI -0.23;-0.04; fat: β=0.53, 95% CI 0.03;1.03; fibroglandular: β=-0.16, 95% CI -0.25;-0.07). We found no associations of alcohol use with any of the tissue measures in premenopausal women. Conclusion: Our findings suggest that both recent and cumulative average alcohol consumption are associated with smaller proportion of stroma and fibroglandular tissue and a greater proportion of fat in postmenopausal women. Future studies are warranted to confirm our findings and to elucidate the underlying biological mechanisms. Citation Format: Lusine Yaghjyan, Rebecca Austin-Datta, Yujing J Heng, Gabrielle Baker, Bernard Rosner, Rulla Tamimi. Associations of alcohol consumption with benign breast tissue composition [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P3-12-19.